Evaluation of In Vivo Antidiarrheal Activities of 80% Methanol Extract and Solvent Fractions of Peels of Colocasia esculenta (Araceae)

Background Diarrhea is the common gastrointestinal disorder accounting for 2.5 billion episodes and 1.5 million deaths annually. Limitations and inaccessibility of currently available medications are the main problem associated with treatment of diarrhea. Hence, medicinal plants are usually preferred to manage diarrhea because they may contain constituents with high activity and fewer side effects. Even though the dose, safety, and efficacy of Colocasia esculenta (L.) Schott are not substantiated scientifically, several societies use it for the treatment of diarrhea. Objective This study was targeted at exploring the in vivo antidiarrheal activities of 80% methanol extract and solvent fractions of peels of Colocasia esculenta (L.) Schott in Swiss albino mice. Methods The plant was collected and extracted with 80% methanol, followed by fractionation with distilled water, chloroform, and diethyl ether. Castor oil-induced diarrhea, enteropooling, and motility tests were used to evaluate antidiarrheal activity. The test groups received graded doses of 100 mg/kg, 200 mg/kg, and 400 mg/kg. Negative controls received 10 ml/kg of 2% Tween 80 while positive controls received loperamide (3 mg/kg) or atropine (5 mg/kg). Results The crude and solvent fractions of the plant extract have induced significant effects in reduction of the number and weight of wet stools at all tested doses. However, delay in onset of diarrhea was observed only at 400 mg/kg (P < 0.001) for both crude extract and solvent fractions. In antienteropooling test, 80% methanol extract and solvent fractions have significantly reduced the weight and volume of intestinal contents, especially at 200 mg/kg and 400 mg/kg. Regarding the antimotility test, the crude extract reduced motility at all tested doses, whereas the solvent fractions reduced intestinal motility mainly at 400 mg/kg (P < 0.001). Conclusion The study has revealed that the 80% methanol extract and solvent fractions of the plant possess antidiarrheal activities supporting the traditional antidiarrheal claims of the plant.


Introduction
Diarrhea is a gastrointestinal disorder characterized by fecal urgency and incontinence.It causes an increase in the frequency, weight, and/or fuid content of the stool [1,2].Diarrhea can be caused by a variety of substances, including medications, poisons, infections, gastrointestinal illnesses, poorly absorbable materials, and issues with infammation and dysmotility in the GI tract [3].It is estimated to be responsible for 3.6% of the world's illness burden and almost 8% of all pediatric deaths.Diarrhea in children under fve causes around 2.5 billion episodes and 1.5 million fatalities worldwide each year.Diarrhea is more common in underdeveloped nations and accounts for over 21 percent of mortality in children under fve [4].Wash and immunizations are the two main ways to avoid diarrhea [5,6].Reversing or preventing dehydration and lowering the morbidity and mortality linked to it are the main objectives of treating diarrhea [7].Supportive therapy, antidiarrheal therapy, and empirical antibiotic therapy are the three primary methods used to treat diarrhea.Te management of diarrhea can beneft greatly from currently prescribed medications, but they are also associated with a number of negative side efects, including bronchospasm, vomiting, intestinal blockage, constipation, dependency, drug resistance, and superinfections [8][9][10].Despite advancements in modern medicine, many individuals, particularly in developing nations, depend on medicinal plants for healthcare, including diarrhea treatment.Traditional healers rely on ancestral knowledge, lacking precise information on dosage, safety, and chemical composition.It is crucial to scientifcally validate the efcacy, safety, and usefulness of active constituents in medicinal plants to ensure their reliable integration into medical practices [11].
Traditional healers have long relied on a variety of medicinal plants with potential antidiarrheal properties, although the scientifc evaluation of their efectiveness remains limited [12].In the context of Ethiopian traditional medicine, numerous traditional remedies have been recorded, including the use of Colocasia esculenta (L.) Schott, commonly known as "Godere," "Yitri," or "Wuhayinkash."Dating back to ancient times, this herb has been known for its healing properties and has been employed to treat various ailments, such as asthma, arthritis, diarrhea, internal hemorrhage, neurological disorders, skin disorders, body ache, cancer, diabetes, and baldness [13,14].However, further scientifc research is needed to fully understand and validate the therapeutic potential of this plant in treating diarrhea and other diseases.Terefore, this study aimed to investigate antidiarrheal activity of 80% methanol extract and solvent fractions of peels of Colocasia esculenta (L.) Schott in Swiss albino mice.Te study's fndings ofer a valuable foundation for the development of innovative antidiarrheal drugs, aiming to overcome limitations associated with existing medications.Furthermore, the scientifc exploration of this plant's antidiarrheal properties not only validates and safeguards traditional wisdom but also fosters cultural diversity and preserves indigenous medicinal practices for the beneft of future generations.Tese fndings serve as a guiding compass for future researchers, students, healthcare providers, and traditional medicine practitioners, facilitating their endeavors in advancing knowledge and improving patient care.

Plant Material.
In March 2021, the peels of Colocasia esculenta were gathered from Deko Kebele, Wonago Woreda, located in the Southern Nation, Nationalities, and Peoples Region of South Ethiopia.Te plant's authenticity was confrmed by a Botanist, Mr. Abiyu Eniyew Molla, afliated with the Department of Biology at the College of Natural and Computational Sciences, University of Gondar.A voucher specimen with the reference number 001/TOT/ 2021 was preserved for future reference purposes.

Experimental Animals.
Te study adhered to established protocols for the care and use of laboratory animals [15].A total of 293 healthy adult Swiss albino mice, both male and female, aged 8-12 weeks and weighing between 20 and 30 g, were used throughout the entire duration of the experiment.Tese mice were procured from the animal unit of the Department of Pharmacology at the University of Gondar.
Te animals were housed in plastic cages, accommodating up to six mice per cage, with wood chip bedding changed every three days.Tey were provided with adequate food and water, and the lighting conditions followed a natural cycle of 12 hours on and 12 hours of.Tis was done to mimic the natural light-dark cycle experienced by the animals in their natural habitat, with light during the day and darkness during the night.Tis approach may help to regulate their circadian rhythms and ensure a more conducive environment for their well-being and normal physiological functioning.Prior to commencing the experiment, the mice were acclimatized to the laboratory environment for a week [16].

Extraction Process.
Te fresh peels of Colocasia esculenta (L.) Schott were carefully washed and cut into small pieces.To protect plant's constituents from sunlight-induced damage, the smaller pieces were dried under shade.Once dried, the plants were coarsely powdered using a mortar and pestle.Approximately 1700 grams of the plant powder was utilized for extraction.Te powder was then subjected to cold maceration in an Erlenmeyer fask, mixed with 80% methanol in a 1 : 5 (w/v) ratio, and left to soak at room temperature for 72 hours with periodic shaking [17].Following the initial extraction, the residue was fltered through muslin cloth and Whatman No. 1 flter paper to separate the liquid from the solid material.To maximize the yield, the remaining solid material underwent two additional extraction cycles using fresh solvent.Te resulting fuid extracts were combined, fltered, and subjected to evaporation at 40 °C using a rotary evaporator.Te concentrated extract was then transferred to a hot air oven to remove any remaining liquid content.Subsequently, the extract underwent a deep-freezing process for 24 hours and was subsequently dried into powder form using a vacuum freeze dryer.Finally, the powder was stored in a refrigerator at 4 °C until it was ready for use [18].Te percentage yield of the extract was found to be 8.23% (w/w).

Fractionation Process.
After confrming the safety and dose-dependent antidiarrheal activity of the crude extract, further fractionation was conducted.A total of 120 g of the crude extract was subjected to successive fractionation using diethyl ether, chloroform, and distilled water.To begin, the crude extract was suspended in 600 mL of distilled water at 2 Evidence-Based Complementary and Alternative Medicine a 1 : 5 ratio.Te suspension was then placed in a separatory funnel, and an equal volume of diethyl ether (600 mL) was added.Te mixture was thoroughly mixed and allowed to settle until distinct layers formed, facilitating the extraction of diethyl ether-soluble constituents.Te diethyl ether fraction (upper layer) was collected in a beaker, and this process was repeated two more times.Following the diethyl ether fraction collection, an equal volume (600 mL) of chloroform was introduced to the remaining aqueous residue and thoroughly mixed.Once a clear layer formed, the chloroform layer at the bottom was carefully collected in a beaker.Tis process was repeated twice more to ensure maximum extraction.Both the diethyl ether and chloroform fractions were subjected to evaporation using a rotary evaporator, and the resulting fractions were stored in a refrigerator at 4 °C.Te remaining aqueous residue was also collected and evaporated at 40 °C using a rotary evaporator.Te resulting residue was then placed in a deep freezer, followed by drying in a lyophilizer to obtain the aqueous fraction.After complete drying, the fraction was stored in an airtight container in the refrigerator until it was ready for use in the subsequent tests [17].

Preliminary Phytochemical Screening
Test.Te 80% methanol extract and the various solvent fractions underwent standard screening tests to determine the presence of specifc phytoconstituents.Tese tests included the screening for favonoids, tannins, anthraquinones/steroids, glycosides, phenols, terpenoids, alkaloids, and saponins.Te purposes were to identify and evaluate the phytochemical composition of the extract and fractions [17].

Acute Oral Toxicity Test.
To assess acute oral toxicity, a total of fve healthy, nonpregnant female Swiss albino mice, aged 8-12 weeks, were employed due to their heightened sensitivity [19].Te mice were acclimatized to the laboratory environment for one week prior to the experiment.Following a fasting period of 4 hours, the frst mouse was administered a dose of 2 g/kg of the 80% methanol extract, in accordance with Organization for Economic Co-operation and Development (OECD) 425 guidelines.Te mouse was then closely monitored for any physical or behavioral changes within a 24-hour period, with particular attention during the initial 4 hours.As the frst mouse displayed no signs of toxicity, the remaining four mice were subsequently treated with the same dose and observed daily for 14 days for any indications of toxicity, such as loss of appetite, hair erection, lacrimation, tremors, convulsions, salivation, diarrhea, mortality, and other adverse efects [20].
2.7.Animal Grouping and Dosing.For the experiment, a healthy adult Swiss albino mouse, both males and females, aged between 8 and 12 weeks and weighing 20− 30 g, was utilized.Te study involved three models and included the 80% methanol extract as well as each of the solvent fractions.Te mice were randomly assigned to fve groups, each consisting of six mice for each model.Group I served as the negative control and received 10 mL/kg of 2% Tween 80 in all models.Group II, the positive control, received loperamide at a dose of 3 mg/kg (10 mL/kg) for the castor oilinduced diarrhea and intestinal fuid accumulation test.Atropine sulfate at a dose of 5 mg/kg was administered intraperitoneally as the positive control for the GI motility test.Groups III, IV, and V were treated with doses of 100 mg/kg, 200 mg/kg, and 400 mg/kg of the Colocasia esculenta (L.) Schott peel extracts, respectively [21,22].

Antidiarrheal Activity on Castor Oil-Induced Diarrhea.
A total of thirty Swiss albino mice, regardless of their sex, were selected and weighed before being randomly assigned to fve groups, with six mice in each group.Te mice underwent an 18-hour fasting period but had access to water.Te dosing and grouping were carried out as described earlier.One hour after administration of the appropriate extract, each mouse was orally administered 0.5 mL of castor oil using a gavage technique.Te mice were then individually placed in metabolic plastic cages lined with nonwetting transparent paper, which was changed whenever the mice defecated.Over a four-hour observation period, various parameters such as the onset of diarrhea, frequency of wet feces, total number of defecations, weight of wet and dry stool, and consistency of feces were recorded and compared to the control group.Te percentage inhibition of diarrhea and defecations was calculated for both the crude extracts and solvent fractions [23][24][25] where MVIC is the mean volume of intestinal content, MVICC is the mean volume of intestinal content of control group, and MVICT is the mean volume of intestinal content of test group.

Gastrointestinal Motility Test by Activated Charcoal
Suspension.A total of thirty mice underwent an 18-hour fasting period, with access to water, and were divided into fve groups of six mice each, as previously mentioned.
Following the animal grouping and dosing procedure, one hour after treatment with the extract and/or controls, the mice were challenged with 0.5 mL of castor oil.After thirty minutes, the mice received 0.5 mL of a 10% charcoal suspension in 2% Tween 80. Tirty minutes after the administration of the charcoal suspension, the mice were sacrifced, and their abdomens were opened to remove the intestine from the pylorus to the cecum.Te intestine was then placed on white paper, and the length of the small intestine and the distance traveled by the charcoal suspension from the pylorus to the cecum were measured using a ruler.Te peristalsis index, representing the percentage of the overall length of the small intestine that the charcoal meal had traversed, was calculated.Finally, the percent inhibition of intestinal transit was determined by comparing the mean distance traveled by the charcoal meal with the controls [28][29][30][31].
Peristalsis index(PI) � distance traveled by the charcoal suspension total length of small intestine * 100, % of inhibition � PI of negative control − PI of drug or extract treated PI of negative control * 100. (4)

In Vivo Antidiarrheal Index (ADI).
Te antidiarrheal index (ADI) was determined through the combination of data obtained from the castor oil-induced diarrhea, enteropooling, and gastrointestinal (GI) motility tests.Te ADI was calculated using a specifc formula, taking into account the results from these tests [22].
where DDT � the delay in defecation time or diarrheal onset (as % of control), GT � the GI motility by charcoal travel reduction (as % of control), and IFA � the reduction in the intestinal fuid accumulation (as % of control

Te Percentage Yield.
A total of 1700 g of plant powder was subjected to extraction, resulting in the production of 140 g of crude extract.Te crude extract (120 g) was further fractionated using solvents of increasing polarity, namely, diethyl ether, chloroform, and distilled water.Te percentage yield was then calculated for each fraction.Te results indicated a percentage yield of 8.23% (140 g) for the crude extract, 46.7% (56 g) for the aqueous fraction, 26.67% (32 g) for the chloroform fraction, and 9.16% (11 g) for the diethyl ether fraction.

Preliminary Phytochemical Screening.
Table 1 shows the outcomes of a preliminary phytochemical screening conducted on both the crude extract and solvent fractions of the plant.Te screening revealed the potential presence of various bioactive compounds, including favonoids, tannins, anthraquinones, steroids, glycosides, phenols, terpenoids, alkaloids, and saponins (Table 1).

Acute Oral Toxicity Test.
Following the oral administration of a single dose of 2000 mg/kg of the hydromethanolic peel extract of Colocasia esculenta (L.) Schott, no deaths or observable signs of toxicity were reported within a 24-hour period.Additionally, throughout a 14-day observation period, no indications of toxicity such as loss of appetite, hair erection, lacrimation, tremors, convulsions, salivation, or diarrhea were observed.Tese fndings suggest that the plant extract possesses a substantial safety margin and can be considered safe for use.

Efects of 80% Methanol Extract of the Peels of Colocasia esculenta (L.) Schott on Castor Oil-Induced Diarrheal Model.
During a four-hour observation period following the administration of castor oil, the three doses (100, 200, and 400 mg/kg) demonstrated a noteworthy decrease in the number of wet stool (P < 0.001), total stool (P < 0.001), weight of wet stool (P < 0.001), and fuid content of stool (P < 0.001) compared to the negative control.However, it was observed that only the dose of 400 mg/kg (P < 0.01) signifcantly delayed the onset of diarrhea.
In contrast, the dose of 400 mg/kg exhibited a signifcant diference in the delay of diarrhea (P < 0.05), total number of wet stool (P < 0.05), and weight of wet stool (P < 0.05) when compared to the 100 mg/kg dose.Te percent inhibition of diarrhea was found to be 44.03%,60.58%, and 65.46% at the doses of 100, 200, and 400 mg/kg, respectively.In comparison, loperamide exhibited a percent inhibition of 78.23%.Similarly, the percent inhibition of defecation was 44.57%, 53.47%, and 63.69% at the doses of 100, 200, and 400 mg/kg, respectively, as presented in Table 2.

Efects of 80% Methanol Extract on Castor Oil-Induced
Enteropooling.Upon dissecting and measuring the weight of small intestinal contents, all three doses (100 mg/kg, 200 mg/kg, and 400 mg/kg) demonstrated a signifcant decrease (P < 0.001) compared to the negative control.Te percent inhibition of small intestinal fuid accumulation, assessed by weight, was 34.5%, 42.3%, and 60.4% at the doses of 100 mg/kg, 200 mg/kg, and 400 mg/kg, respectively.Loperamide exhibited a percent inhibition of 65.7% in the same regard.
In terms of the volume of small intestinal contents, all three doses showed a signifcant diference (P < 0.001) compared to the negative control.Additionally, the doses of 200 mg/kg and 400 mg/kg demonstrated a signifcant difference (P < 0.05) in comparison with the 100 mg/kg dose.Te percent inhibition of small intestinal secretion, measured by volume, was 41%, 57%, and 66.4% at the doses of 100 mg/kg, 200 mg/kg, and 400 mg/kg, respectively, while the positive control exhibited a percent inhibition of 72.4%, as illustrated in Table 3.

Efects of 80% Methanol Extract on Castor Oil-Induced
Gastrointestinal Motility.All doses of the crude extract from the plant exhibited a signifcant reduction (P < 0.001) in the peristalsis index compared to the negative controls.Furthermore, the dose of 400 mg/kg demonstrated a signifcant decrease (P < 0.001) in the peristalsis index when compared to the 100 mg/kg dose.Te percent inhibition of intestinal transit of the charcoal meal through the gastrointestinal tract (GIT) was found to be 18.2%, 22.3%, and 38.9% at the doses of 100 mg/kg, 200 mg/kg, and 400 mg/kg, respectively.In comparison, the positive control, atropine (5 mg/kg), exhibited a percent inhibition of 64.4%, as depicted in Table 4.  Evidence-Based Complementary and Alternative Medicine Evidence-Based Complementary and Alternative Medicine

Efects of Solvent Fractions of the Plant on Castor
Oil-Induced Diarrheal Model.In the castor oil-induced diarrheal model, all three fractions signifcantly reduced the total number of wet stool and total stool at doses of 100 mg/kg (P < 0.001), 200 mg/kg (P < 0.001), and 400 mg/kg (P < 0.001) when compared to the negative control.Both the aqueous and chloroform fractions signifcantly reduced the weight of wet stool at 200 mg/kg (P < 0.001) and 400 mg/kg (P < 0.001), while the diethyl ether fraction only showed a reduction at 400 mg/kg (P < 0.001) compared to the negative control.All solvent fractions produced a signifcant delay in the onset of diarrhea and reduction in the fuid content of the stool, but only at a dose of 400 mg/kg compared to the negative control, with varying levels of signifcance.Te percent inhibition of diarrhea for the aqueous fraction was 31.67%,44.86%, and 55% at doses of 100 mg/kg, 200 mg/kg, and 400 mg/kg, respectively, while the standard drug loperamide exhibited a percent inhibition of 70.69% (Table 5).Te chloroform fraction did not exhibit signifcant activity at doses of 100 mg/kg and 200 mg/kg in terms of the delay in onset of diarrhea and fuid content of the stool compared to the negative control.However, it did show a signifcant diference in all tested parameters compared to the positive control.Te percent inhibition of diarrhea by the chloroform fraction was 30.75%, 40.85%, and 57.73% at doses of 100 mg/kg, 200 mg/kg, and 400 mg/kg, respectively.Similarly, the diethyl ether fraction did not demonstrate signifcant activity at doses of 100 mg/kg and 200 mg/kg in relation to the weight of wet stool, fuid content of the stool, and delay in onset of diarrhea compared to the negative control.Te percentage inhibition of diarrhea by the diethyl ether fraction was 27.5%, 37.23%, and 55.94% at doses of 100 mg/kg, 200 mg/kg, and 400 mg/kg, respectively (Table 5).When comparing the efcacy among the doses of the solvent fractions, the highest dose of 400 mg/kg exhibited signifcant activity in all tested parameters compared to doses of 100 mg/kg and 200 mg/kg.

Efects of Solvent Fractions on Castor Oil-Induced
Enteropooling Model.Te aqueous fraction derived from the peels of Colocasia esculenta (L.) Schott exhibited signifcant reductions in both the mean weight and volume of intestinal contents at doses of 100 mg/kg (P < 0.05 for volume, P < 0.01 for weight), 200 mg/kg (P < 0.001), and 400 mg/kg (P < 0.001) compared to the negative control.Te percentage inhibition of the weight of intestinal content was 29.68%, 45.36%, and 58.9% at doses of 100 mg/kg, 200 mg/ kg, and 400 mg/kg, respectively.Similarly, the percentage inhibition by volume was 31.2%, 49.2%, and 61.7% at the corresponding doses.Moreover, the dose of 400 mg/kg demonstrated a signifcant diference compared to the 100 mg/kg dose of both the aqueous fraction and chloroform fraction in terms of the weight and volume of intestinal contents, with diferent levels of signifcance, as presented in Table 6 (Figure 1).
Similarly, the chloroform fraction also exhibited signifcant reductions in the weight and volume of intestinal contents at all doses: 100 mg/kg (P < 0.01 for weight),   Evidence-Based Complementary and Alternative Medicine 9 200 mg/kg (P < 0.001 for weight, P < 0.01 for volume), and 400 mg/kg (P < 0.001) compared to the negative control.However, the 100 mg/kg dose did not show a signifcant efect on the volume of intestinal contents.Te percent inhibition of weight and volume of intestinal contents was 22.2%, 39.6%, and 53.78% for weight, and 22.38%, 41.22%, and 52.7% for volume at doses of 100 mg/kg, 200 mg/kg, and 400 mg/kg, respectively (Figure 1).Similarly, the diethyl ether fraction also signifcantly reduced the weight and volume of intestinal contents (P < 0.001) compared to the negative control.Te percent inhibition of weight and volume of intestinal contents was 26.7%, 41.2%, and 60.2% for weight, and 27%, 43%, and 62.6% for volume at doses of 100 mg/kg, 200 mg/kg, and 400 mg/kg, respectively (Table 6) (Figure 1).Te highest dose of 400 mg/kg of all solvent fractions demonstrated signifcant efects on the weight and volume of intestinal contents compared to the 100 mg/kg dose, with diferent levels of signifcance.Te percent inhibition by both weight and volume decreased from 400 mg/kg to 100 mg/kg in all solvent fractions (Figure 1).

Efects of Solvent Fractions on Castor
Oil-Induced Gastrointestinal Motility.Te aqueous fraction of the plant extract signifcantly inhibited intestinal transit of charcoal meal at a dose of 400 mg/kg (P < 0.001) compared to the negative control, while the doses of 100 mg/kg and 200 mg/ kg did not show a signifcant efect on intestinal motility.All three doses of the aqueous fraction demonstrated a signifcant diference (P < 0.001) in intestinal motility compared to for doses of 100 mg/kg, 200 mg/kg, and 400 mg/kg, respectively, whereas atropine exhibited a 65% inhibition (Table 7).
Likewise, the diethyl ether fraction signifcantly afected intestinal propulsion of charcoal meal at a dose of 400 mg/kg (P < 0.01) compared to the negative control.Te percent inhibition of intestinal transit of charcoal meal was 6.8%, 12.6%, and 27% for doses of 100 mg/kg, 200 mg/kg, and 400 mg/kg, respectively.
3.10.In Vivo Antidiarrheal Index.Te in vivo antidiarrheal indexes of both crude extract and solvent fractions of peels of Colocasia esculenta (L.) Schott have exhibited a variation among diferent doses the plant extract as shown in Figure 2.

Discussion
Te antidiarrheal properties of the peels of Colocasia esculenta (L.) Schott, a herbaceous plant traditionally used for managing diarrhea in various countries, were investigated in this study.Tree diferent diarrhea models, including castor oil-induced diarrhea, enteropooling, and gastrointestinal motility tests, were conducted on Swiss albino mice to evaluate the claimed antidiarrheal activity.
In this study, the induction of diarrhea was achieved using castor oil, which contains ricinoleic acid as its active metabolite.Ricinoleic acid is known to stimulate the secretory process and increase intestinal motility by causing irritation and infammation of the intestinal mucosal linings.Tis infammation leads to the release of various infammatory mediators, including prostaglandin (PGE-2), which can afect ion channels and disrupt normal physiological processes.Consequently, these infammatory mediators, particularly PGE-2 and nitric oxide (NO), can inhibit glucose absorption, trigger infammation of the intestinal mucosa, cause contraction of the intestinal smooth muscle, and disrupt ion channels, such as Na + -K + ATPase.Tese efects collectively contribute to an increase in intestinal secretion and motility [32,33].
Previous studies [23,27,30] have indicated that various plant constituents such as saponins, favonoids, alkaloids, terpenoids, phenols, anthraquinones, steroids, glycosides, and tannins are believed to be responsible for the antidiarrheal efects observed in medicinal plants.In the present study, phytochemical screening was conducted on the peels of Colocasia esculenta (L.) Schott, and it revealed the presence of these bioactive compounds in both the 80% methanol extract and solvent fractions.Tis fnding supports the claim of the plant extract's antidiarrheal properties, although the specifc constituents responsible for this efect have yet to be identifed.
In the castor oil-induced diarrheal model, the frequency and weight of wet stool were given particular attention as they are commonly associated with diarrhea.Te 80% methanol extract of plant's peels demonstrated signifcant activity at all tested doses in reducing the frequency and weight of wet stool.Notably, the dose of 400 mg/kg exhibited the highest inhibition of diarrheal stool (65.46%), indicating an increased efectiveness of the extract with higher doses.
Te crude extract of the peels of Colocasia esculenta (L.) Schott demonstrated superior activity in inhibiting diarrheal stool compared to total defecation when compared to the negative control.Additionally, the crude extract was able to reduce the fuid content of the stool at all tested doses, indicating its ability to enhance colonic fuid absorption.
Regarding the delay of diarrhea onset, the dose of 400 mg/kg exhibited better activity compared to the standard drug loperamide.Tis may be attributed to the extract's essential absorptive properties and its ability to block chloride channels.However, the doses of 100 mg/kg and 200 mg/kg of the plant extract did not show a signifcant efect on delaying diarrhea.Tis could be due to the lower concentrations of phytochemicals present in these doses, which might not be sufcient to produce the desired therapeutic efect.
Overall, the crude extract of the peels of Colocasia esculenta (L.) Schott reduced all tested parameters at different doses, although the efcacy varied among the doses.Te signifcant efects of the crude extract can be attributed to the presence of various phytoconstituents, particularly alkaloids, tannins, favonoids, phenols, and terpenoids.Tese compounds inhibit prostaglandin (PG) secretion, fuid, and electrolyte secretion, while promoting absorption through diferent mechanisms.Furthermore, the plant's anti-infammatory efects observed in in vivo studies could contribute to its enhanced efcacy [34].
Consistent with the previous fndings, the solvent fractions of the plant exhibited a signifcant reduction in the total number of wet stool and the number of defecations at all tested doses.However, the solvent fractions only reduced the weight of wet stool at doses of 200 mg/kg and 400 mg/kg compared to the negative control.Regarding the onset of diarrhea and fuid content of the stool, all fractions demonstrated a signifcant reduction only at the dose of 400 mg/ kg, indicating a dose-dependent efect of these parameters.
Furthermore, in the castor oil-induced diarrheal model, the 80% methanol extract was found to be more efective than the solvent fractions in terms of all tested parameters.Tis diference in efectiveness could be attributed to variations in the availability and concentrations of phytoconstituents among the diferent fractions.Among the solvent fractions, the aqueous fraction exhibited the highest efectiveness in inhibiting diarrhea, while the diethyl ether fraction was the least efective.However, the aqueous fraction showed the highest efectiveness in inhibiting defecations, and the chloroform fraction was the most effective in delaying the onset of diarrhea.

Evidence-Based Complementary and Alternative Medicine
Te less efective performance of the aqueous fraction in delaying diarrhea could be due to the absence of favonoids, which play a crucial role in inhibiting prostaglandin (PG) release and gastrointestinal motility.Additionally, the available constituents in the aqueous fraction may not reach therapeutic concentrations, thereby limiting their intended efects.Conversely, the high efectiveness of the chloroform fraction in delaying diarrhea could be attributed to the presence of constituents such as alkaloids, favonoids, and phenols, which are known to exhibit the required activity.Phenols can reduce intestinal secretion and enhance absorption through their antioxidant, anti-infammatory, and antibacterial properties.Tannins cystic fbrosis transmembrane conductance regulator (CFTR) and calcium-activated chloride channel (CaCC), thereby decreasing chloride secretions.Flavonoids have the potential to inhibit cyclooxygenase-1 (COX-1), cyclooxygenase-2 (COX-2), and lipoxygenase (LOX), thus reducing prostaglandin synthesis.Tey can also reduce intestinal motility by activating α2-adrenergic receptors, leading to sympathetic nerve activation and subsequent reduction in intestinal contractions.All of these factors contribute to the delay in the onset of diarrhea, decrease in fuid content of the stool, and reduce the frequency and weight of stool [35][36][37][38].
In castor oil-induced enteropooling model, the crude extract at all tested doses exhibited signifcant reductions in the mean weight and volume of intestinal contents compared to the negative control.Notably, the dose of 400 mg/kg demonstrated the highest percentage of inhibition (60.4% for weight and 66.4% for volume) among all the doses tested.Tis suggests a clear dose-response relationship for the plant extract in this context.
Regarding the antienteropooling efects of the solvent fractions, all fractions demonstrated signifcant reductions in both the average weight and volume of intestinal contents, although there were variations among the diferent doses.12 Evidence-Based Complementary and Alternative Medicine However, the chloroform fraction exhibited the least activity in reducing intestinal fuid accumulation compared to the other fractions.Tis could be attributed to the absence of metabolites such as saponins, tannins, glycosides, and steroids, which are known to play a crucial role in antisecretory activities.
Te crude extract of the plant showed signifcant antienteropooling efects at all tested doses, with the highest inhibition observed at the dose of 400 mg/kg.Similarly, the solvent fractions demonstrated reductions in intestinal fuid accumulation, the chloroform fraction was comparatively less efective, possibly due to the absence of certain metabolites [35,39].Te presence of tannins and steroids in the diethyl ether fraction could contribute to its higher efectiveness compared to the chloroform fraction.Steroids are known to induce antienteropooling efects through their anti-infammatory properties, inhibiting the release of prostaglandin E-2 (PGE-2) and prostacyclin (PGI-2) from macrophages.On the other hand, tannins can inhibit secretion and reduce fuid accumulation by modulating the activity of cystic fbrosis transmembrane conductance regulator (CFTR) and calcium-activated chloride channel (CaCC) [40].Other phytoconstituents present in the plant extract, such as alkaloids, glycosides, and anthraquinones, may also contribute to the antisecretory efects observed.Tese compounds have been reported to possess antibacterial, antifungal, anti-infammatory, and antioxidant activities, which can collectively contribute to the inhibition of fuid secretion and accumulation in the intestines [38,41].
Te crude extract of Colocasia esculenta (L.) Schott, which contains a combination of phytochemicals, demonstrated superior efcacy compared to the solvent fractions in reducing intestinal fuid accumulation and motility.Te aqueous fraction exhibited signifcant antisecretory efects, while the chloroform fraction showed the least efect in both parameters.Te crude extract displayed a clear doseresponse relationship, with higher doses being more efective in inducing antimotility efects, although the percentage inhibition of intestinal propulsion was lower compared to the standard drug atropine sulfate.
Among the solvent fractions, the chloroform fraction exhibited the highest activity in reducing intestinal motility.Tis may be attributed to the presence of intermediate polarity phytochemicals such as favonoids, alkaloids, and phenols, which are known to impact peristalsis.However, it is important to note that the overall activity of the solvent fractions in inhibiting intestinal motility was lower than the standard drug.Te presence and concentration of bioactive compounds, including tannins, terpenoids, favonoids, phenols, and alkaloids, likely contribute to the regulation of peristalsis, with tannins reducing calcium infux and favonoids activating α2adrenergic receptors, resulting in decreased motility and prolonged transit time [35,38].In previous studies, it has been indicated that the presence of favonoids, alkaloids, and phenols may contribute to antidiarrheal efects.Tese efects are believed to occur through various mechanisms, including the inhibition of arachidonic acid metabolism, promotion of fuid and electrolyte absorption, inhibition of secretion, and reduction of intestinal motility [42].
Hence, those abovementioned notions suggested another possible antidiarrheal mechanism of the plant extract that could be inhibition of intestinal motility via blockage of Ca 2+ channel, inhibition of PG and NO release, hampering of Ach and 5-HT release, and/or preventing infammation of intestinal mucosa [43].Hence, a decrease in intestinal motility allows more time for absorption of intestinal contents.
Te assessment of antidiarrheal activity in medicinal plants often relies on in vivo antidiarrheal indices (ADI), which consider multiple parameters, such as the onset of diarrhea, mean weight of wet stools, and distance traveled by charcoal suspension [30].In this study, the crude extracts and diethyl ether fraction displayed the highest and lowest ADI, respectively, across all tested doses.Te increase in ADI indicates an enhanced antidiarrheal activity.Tese fndings suggest that the crude extract and solvent fractions of Colocasia esculenta (L.) Schott peels possess signifcant antidiarrheal potential, as evidenced by the notable reduction in key parameters associated with diarrhea.
Overall, the in vivo antidiarrheal activities of the 80% methanol extract and solvent fractions of Colocasia esculenta (L.) Schott peels indicate their potential use in managing diarrhea, as demonstrated in the conducted models using Swiss albino mice.Tese fndings highlight the efectiveness of the plant extract as a possible antidiarrheal agent.By considering the diferent aspects of diarrhea and its associated symptoms, the study provides evidence supporting the utilization of Colocasia esculenta (L.) Schott as a natural remedy for diarrhea management.

Conclusion
Te current study demonstrated that both the crude extract and solvent fractions of Colocasia esculenta (L.) Schott peel extracts possess signifcant antidiarrheal activity, likely attributed to their ability to absorb fuids, reduce secretion, and moderately inhibit motility.Te evaluation of the in vivo antidiarrheal index indicated that the plant extracts exhibited moderate efects across all tested parameters, with increasing doses showing more pronounced activity.Furthermore, the phytochemical analysis of the plant extracts revealed the presence of various bioactive compounds responsible for the observed antidiarrheal efects.Tese fndings provide scientifc evidence supporting the traditional use of Colocasia esculenta (L.) Schott in the treatment of diarrhea.

Figure 2 :
Figure 2: In vivo antidiarrheal index of crude extract and solvent fractions of peels of Colocasia esculenta (L) Schott. .
Prior to commencing the experimental activities, the research proposal was submitted to and approved by the Department Graduate Committee of Pharmacology at the University of Gondar.Additionally, ethical clearance was obtained from the same department, with a reference number of Sop4/103/2013.Te handling and care of the experimental animals adhered to both local and international guidelines regarding the use, care, and welfare of laboratory animals.2.10.Statistical Analysis.Te results were presented as the mean ± standard error of the mean (SEM) and were subjected to statistical analysis using SPSS version 26.0.Group comparisons were evaluated using one-way analysis of variance (ANOVA), followed by a post hoc Tukey test.Statistical signifcance was considered for probability values below 0.05.Te processed data were summarized and presented in tabular form.
). DDT � Onset of diarrhea in a minute of the ( test − negative control) group Onset of diarrhea in a minute of the negative control group * 100, GMT � Distance traveled by the charcoal marker of the ( negative control test) group Distance traveled by the charcoal marker in the negative control group * 100, IFA � Mean weight of wet stools of (the negative control − treated) group Mean weight of wet stools of negative control group * 100.(6) 4 Evidence-Based Complementary and Alternative Medicine 2.9.Ethical Clearance.2.11.Data Quality Assurance.Data were compiled, cleared, coded, and checked for completeness and accuracy before entering into SPSS version 26.0.Mice were assigned to each group through random selection to reduce bias.

Table 2 :
Efects of the 80% methanol extract on castor oil-induced diarrheal model in mice.

Table 3 :
Efects of the 80% methanol extract on castor oil-induced enteropooling model in mice.

Table 4 :
Efects of the 80% methanol extract on castor oil-induced gastrointestinal motility model in mice.

Table 5 :
Efects of solvent fractions on castor oil-induced diarrheal model.

Table 6 :
10 efects of solvent fractions on castor oil-induced enteropooling model in mice.Values are expressed by mean ± SEM (n � 6); analysis was performed by one-way ANOVA, followed by post hoc Tukey.Comparisons were made between diferent groups of study; a compared with negative control; b compared with positive control (loperamide 3 mg/kg).cComparedwith 100 mg/kg AF; d compared with 200 mg/kg AF; e compared with 400 mg/kg AF; f compared with 100 mg/kg CHF; g compared with 200 mg/kg CHF; h compared with 400 mg/kg CHF; i compared with 100 mg/kg DEEF; j compared with 200 mg/kg DEEF; k compared with 400 mg/kg DEEF; 1 P < 0.05, 2 P < 0.01, and 3 P < 0.001.AF, aqueous fraction; CHF, chloroform fraction; DEEF, diethyl ether fraction.NB: negative control 2% Tween 80 for all fractions.Te efect of 80% methanol extract and solvent fractions of peels of Colocasia esculenta (L.) Schott on the percent inhibition in a volume of intestinal content.Group 1: negative control, group 2: positive control, group 3: 100 mg/kg, group 4: 200 mg/kg, and group 5: 400 mg/kg.10Evidence-BasedComplementary and Alternative Medicine the standard drug, atropine sulfate.Te percentage inhibition of intestinal propulsion was 9%, 16.8%, and 31.8%

Table 7 :
Te efects of the solvent fractions on castor oil-induced gastrointestinal motility model in mice.Data are expressed by mean ± SEM (n � 6); analysis was conducted by one-way ANOVA, followed by post hoc Tukey.Comparisons were made between diferent groups of study; a compared with negative control; b compared with positive control (atropine 5 mg/kg).c Compared with 100 mg/kg AF; d compared with 200 mg/kg AF; e compared with 400 mg/kg AF; f compared with 100 mg/kg CHF; g compared with 200 mg/kg CHF; h compared with 400 mg/kg CHF; compared with 200 mg/kg DEEF; k compared with 400 mg/kg DEEF; 1 P < 0.05, 2 P < 0.01, and 3 P < 0.001.AF, aqueous fraction; CHF, chloroform fraction; DEEF, diethyl ether fraction.NB: negative control 2% Tween 80 for all fractions.
i compared with 100 mg/kg DEEF; j